Martínez-Costa Oscar H, Sánchez-Martínez Cristina, Sánchez Valentina, Aragón Juan J
Departamento de Bioquímica de la UAM, Instituto de Investigaciones Biomédicas Alberto Sols UAM-CSIC, Facultad de Medicina de la Universidad Autónoma de Madrid, Arzobispo Morcillo 4, 28029 Madrid, Spain.
FEBS Lett. 2007 Jun 26;581(16):3033-8. doi: 10.1016/j.febslet.2007.05.059. Epub 2007 May 29.
Two phosphofructokinase (PFK) chimeras were constructed by exchanging the N- and C-terminal halves of the mammalian M- and C-type isozymes, to investigate the contribution of each terminus to the catalytic site and the fructose-2,6-P(2)/fructose-1,6-P(2) allosteric site. The homogeneously-purified chimeric enzymes organized into tetramers, and exhibited kinetic properties for fructose-6-P and MgATP similar to those of the native enzyme that furnished the N-terminal domain in each case, whereas their fructose-2,6-P(2) activatory characteristics coincided with those of the isozyme that provided the C-terminal half. This reflected the role of each domain in the formation of the corresponding binding site. Grafting the N-terminus of PFK-M onto the C-terminus of the fructose-1,6-P(2) insensitive PFK-C restored transduction of this signal to the catalytic site, which significance is also discussed.
通过交换哺乳动物M型和C型同工酶的N端和C端结构域构建了两种磷酸果糖激酶（PFK）嵌合体，以研究每个结构域对催化位点以及果糖-2,6-二磷酸/果糖-1,6-二磷酸变构位点的贡献。均一纯化的嵌合酶组装成四聚体，对果糖-6-磷酸和MgATP表现出的动力学特性类似于在每种情况下提供N端结构域的天然酶，而它们的果糖-2,6-二磷酸激活特性与提供C端结构域的同工酶一致。这反映了每个结构域在相应结合位点形成中的作用。将PFK-M的N端嫁接到对果糖-1,6-二磷酸不敏感的PFK-C的C端，恢复了该信号向催化位点的转导，其意义也进行了讨论。
